Our goal is to develop inhibitors against a novel and unexploited target for the treatment of AIDS. The human immunodeficiency virus type 1 (HlV-1), the causative agent of AIDS, is a complex retrovirus that encodes six regulatory proteins, including Vif that is essential for viral replication in vivo. Despite remarkable medical advances, HlV-1 infections continue to rise throughout the world. Current anti-HIV-1 agents target mainly HlV-1 reverse transcriptase or protease. However, inhibitors used to target these enzymes have given rise to viral resistance and related toxic effects, creating a need for more potent and less toxic therapies against other viral targets. HIV-1 is a complex retrovirus that encodes 6 regulatory proteins, among which Vif is essential for in vivo viral replication. HIV-1 Vif protein targets an innate antiviral human DNA-editing enzyme, AP0BEC3G (A3G), which inhibits replication of retroviruses. Since HiV-1 Vif has no known cellular homologs, this protein represents an extremely attractive; yet unrealized, target for antiviral intervention. Over the last few years, in collaboration with Dr. Stevenson (Project 2) and Core B, we developed and employed high throughput screening (HTS) assays to identify small molecules that antagonize HIV-1 Vif function. We focused our efforts to characterize one of these lead molecules, RN-18, which inhibits HlV-1 replication only in the presence of A3G. Mechanistic studies revealed that RN-18 enhanced Vif degradation only in the presence of A3G, increased A3G incorporation into virions leading to less infectious viruses, and enhanced cytidine deamination of the viral genome. These studies provide the first evidence that the HIV-1 Vif-A3G axis is a valid target for developing small molecule-based new therapies for AIDS or for enhancing innate immunity against viruses. Based on our preliminary success to develop Vif antagonists as a new class of AIDS therapeutics, we plan to employ highly innovative, collaborative, and multidisciplinary approaches to accomplish our proposed goals. Our project has the following specific aims: Specific Aim 1: Structure activity relationship studies and validation of lead compounds. Specific Aim 2: Structure activity relationship studies, lead optimization, and validation of unexplored scaffolds of Vif antagonists. Specific Aim 3: (i) Mechanism of Vif inhibition. Experiments will be performed to understand the mechanism of Vif inhibitors at molecular level, (ii) In vivo evaluation of Vif inhibitors. During the course of pharmacology studies (Project 3 and core B), we will measure the drug concentrations in plasma and CNS of drug treated animals.